Reduction of friction by recombinant human proteoglycan 4 in IL-1α stimulated bovine cartilage explants.

A boundary lubricant attaches and protects sliding bearing surfaces by preventing interlocking asperity-asperity contact. Proteoglycan-4 (PRG4) is a boundary lubricant found in the synovial fluid that provides chondroprotection to articular surfaces. Inflammation of the diarthrodial joint modulates local PRG4 concentration. Thus, we measured the effects of inflammation, with Interleukin-1α (IL-1α) incubation, upon boundary lubrication and PRG4 expression in bovine cartilage explants. We further aimed to determine whether the addition of exogenous human recombinant PRG4 (rhPRG4) could mitigate the effects of inflammation on boundary lubrication and PRG4 expression in vitro. Cartilage explants, following a 7 day incubation with IL-1α, were tested in a disc-on-disc configuration using either rhPRG4 or saline (PBS control) as a lubricant. Following mechanical testing, explants were studied immunohistochemically or underwent RNA extraction for real-time polymerase chain reaction (RT-PCR). We found that static coefficient of friction (COF) significantly decreased to 0.14 ± 0.065 from 0.21 ± 0.059 (p = 0.014) in IL-1α stimulated explants lubricated with rhPRG4, as compared to PBS. PRG4 expression was significantly up regulated from 30.8 ± 19 copies in control explants lubricated with PBS to 3330 ± 1760 copies in control explants lubricated with rhPRG4 (p < 0.001). Explants stimulated with IL-1α displayed no increase in PRG4 expression upon lubrication with rhPRG4, but with PBS as the lubricant, IL-1α stimulation significantly increased PRG4 expression compared to the control condition from 30.8 ± 19 copies to 401 ± 340 copies (p = 0.015). Overall, these data suggest that exogenous rhPRG4 may provide a therapeutic option for reducing friction in transient inflammatory conditions and increasing PRG4 expression. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:580-589, 2017.

Arthroscopic irrigation of the bovine stifle joint increases cartilage surface friction and decreases superficial zone lubricin.

The purpose of this study was to determine the effects of arthroscopic irrigation on cartilage superficial zone lubricin and surface friction. Arthroscopic partial meniscectomy is one of the most commonly performed orthopedic surgeries in the United States, but rates of osteoarthritis progression following this procedure are high. The effect of arthroscopic irrigation on articular surface lubrication has not been previously considered as a contributing factor in outcomes after arthroscopy. Fourteen bovine stifle joints were randomized to receive arthroscopic irrigation (n=7) or no treatment (n=7). Full-thickness osteochondral explants from these joints underwent friction testing to measure static and dynamic coefficients of friction. Following mechanical testing, samples were fixed and stained for lubricin. Percent integrated density, a measure of the amount of lubricin in the superficial zone (0-100µm depth), was determined. Static and dynamic coefficients of friction were found to be significantly greater in arthroscopy specimens compared to controls (p=0.02 and p<0.001, respectively). Percent integrated density of lubricin in the superficial zone was significantly lower in arthroscopy specimens compared to controls (p<0.001).

Anti-lubricin monoclonal antibodies created using lubricin-knockout mice immunodetect lubricin in several species and in patients with healthy and diseased joints.

Lubricin, encoded by the gene PRG4, is the principal lubricant in articulating joints. We immunized mice genetically deficient for lubricin (Prg4-/-) with purified human lubricin, and generated several mAbs. We determined each mAb's binding epitope, sensitivity, and specificity using biologic samples and recombinant lubricin sub-domains, and we also developed a competition ELISA assay to measure lubricin in synovial fluid and blood. We found the mAbs all recognized epitopes containing O-linked oligosaccharides conjugated to the peptide motif KEPAPTTT. By western blot, the mAbs detected lubricin in 1 µl of synovial fluid from several animal species, including human. The mAbs were specific for lubricin since they did not cross-react with other synovial fluid constituents from patients with camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP), who genetically lack this protein. The competition ELISA detected lubricin in blood samples from healthy individuals but not from patients with CACP, indicating blood can be used in a diagnostic test for patients suspected of having CACP. Lubricin epitopes in blood do not represent degradation fragments from synovial fluid. Therefore, although blood lubricin levels did not differentiate patients with inflammatory joint disease from healthy controls, epitope-specific anti-lubricin mAbs could be useful for monitoring disease activity in synovial fluid.